Synthetic strategy of nonreducing iterative polyketide Synthases and the origin of the classical "Starter-Unit effect"
Synthetic strategy of nonreducing iterative polyketide Synthases and the origin of the classical "Starter-Unit effect"
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DOI：
10.1002/cbic.200700702
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发表时间：
2008-05-05
期刊：
影响因子：
3.2
通讯作者：
Townsend, Craig A.
中科院分区：
文献类型：
--
作者：
Crawford, Jason M.;Vagstad, Anna L.;Townsend, Craig A.
从真菌中分离的聚酮化合物通常来源于“迭代”I型（多结构域）聚酮化合物脱氢酶（PKS），其中各个催化结构域融合成单个大蛋白质，并在给定产物的“程序化”合成中重复使用固定次数[1，2]。迭代I型酶的最著名的例子是酵母和动物脂肪酸脱氢酶（FAS）[3]。动物脂肪酸生物合成由乙酰辅酶A启动，乙酰辅酶A通过双功能酰基转移酶丙二酰辅酶A：酰基载体蛋白（ACP）转酰酶（MAT）进入酶中，MAT主要穿梭丙二酰辅酶A单位进行每次连续的两碳同源化，产生棕榈酸[4]。另一方面，在真菌中，α6β6异十二聚体FAS复合物具有用于起始单元引入的特异性乙酰基转酰酶[5]。在脂肪酸和真菌天然产物的经典前体掺入实验中，通常观察到乙酸起始单元将比分子中的其余标记位点具有更高的来自[14 C]-乙酸的放射性标记的特异性掺入，这与每个链延伸的乙酰辅酶A向丙二酰辅酶A的中间转化一致[6]。相反，如果检查[14 C]-丙二酸盐的互补掺入，则起始碳的标记通常会明显低于聚酮化合物代谢物的其余部分。这种一般模式被称为“起始单元效应”[7]。应用UMA算法选择PksA（GenBank登录号AY 371490）中的结构域间切割位点，PksA是真菌毒素黄曲霉毒素生物合成的迭代I型PKS，揭示了两个先前未识别但明确解析的结构域[8]。最近确定大的N-末端结构域是起始单元：ACP转酰酶（SAT），其选择性地将己酰基起始单元引入PksA ACP上以引发去甲甲异烟酸（1）生物合成[9]。基因组数据库的搜索显示，SAT域广泛存在于已知的和明显的非还原性真菌PKS。PksA在真菌PKS中是不寻常的，因为罕见的己酰基起始单元由FAS亚基HexA和HexB的专用酵母样对提供[10，11]。然而，绝大多数真菌聚酮化合物由乙酰辅酶A和丙二酰辅酶A合成。这些酶不需要专门的设备来制备起始单元，当这些基本的构建单元已经在细胞中可用时。在本文中，我们比较了四个相关的非还原
Polyketides isolated from fungi are typically derived from “iterative” Type I (multidomainal) polyketide synthases (PKSs), where individual catalytic domains are fused into single large proteins, and reused a fixed number of times in the “programmed” synthesis of a given product [1, 2]. The best known examples of iterative Type I enzymes are the yeast and animal fatty acid synthases (FASs)[3]. Animal fatty acid biosynthesis is initiated by acetyl-CoA, which is brought onto the enzyme by a bifunctional acyl transferase, malonyl-CoA: acyl-carrier protein (ACP) transacylase (MAT), which primarily shuttles units of malonyl-CoA for each successive two-carbon homologation leading to palmitate [4]. On the other hand, in fungi an α6β6 heterododecameric FAS complex harbors a specific acetyl transacylase for starter unit introduction [5]. In classical precursor incorporation experiments with fatty acids and fungal natural products it was commonly observed that an acetate starter unit would bear a higher specific incorporation of radiolabel from [14C]-acetate than the rest of the labeled sites in the molecule, in keeping with the intermediary conversion of acetyl-CoA to malonyl-CoA for each chain extension [6]. Conversely, if the complementary incorporation of [14C]-malonate were examined, the starter carbons would often be distinguishably less labeled than the remaining portions of the polyketide metabolite. This general pattern came to be known as the “starter unit effect”[7].Application of the UMA algorithm to select interdomainal cut sites in PksA (GenBank accession no. AY371490), the iterative Type I PKS central to the biosynthesis of the mycotoxin aflatoxin, revealed two previously unrecognized, but clearly resolved domains [8]. The large N-terminal domain was recently established to be a starter unit: ACP transacylase (SAT) that selectively introduced hexanoyl starter units onto the PksA ACP to prime norsolorinic acid (1) biosynthesis [9]. Search of genome databases revealed that SAT domains were widespread among known and apparent nonreducing fungal PKSs. PksA is unusual among fungal PKSs in that the rare hexanoyl starter unit is supplied by a dedicated yeast-like pair of FAS subunits, HexA and HexB [10, 11]. The vast majority of fungal polyketides, however, are synthesized from acetyl-CoA and malonyl-CoA. These enzymes require no specialized apparatus to prepare a starter unit when these rudimentary building blocks are already available in the cell. In this paper we compare four related nonreducing